Separation and analysis of DNA sequence reaction products by capillary gel electrophoresis.
This paper demonstrates the potential of capillary gel electrophoresis with laser induced fluorescence detection as a tool for DNA sequence determination. Both synthetic oligonucleotides and single-stranded phage DNA were utilized as templates in the standard chain termination procedure. Primer molecules were tagged at the 5' end with the fluorescent dye, JOE. First, baseline resolution of a dA extended primer from 18 to 81 bases long, a total of 64 fragments, was observed. A second synthetic template was designed to yield alternating stretches of dA and dT extensions of the primer. Thirdly, the sequence reaction products from a synthetic oligonucleotide template containing all four bases was analyzed in four independent runs, one for each of the four base-specific reactions. In all cases, the expected number and patterns of peaks were observed by capillary gel electrophoretic analysis. Finally, separation of sequence reaction products generated with single-strand M13mp18 phage DNA as template exhibited baseline resolution of fragments differing in length by a single nucleotide and from 18 to greater than 330 bases total length.
Cohen
A S
AS
Barnett Institute, Northeastern University, Boston, MA 02115.
Najarian
D R
DR
Karger
B L
BL
D001483:Base Sequence; D004247:DNA / Q000032:analysis* / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods*; D008969:Molecular Sequence Data
D004247:DNA